This invention relates to apparatus and methods for analyzing cytoplasmic components of single cells in large heterogeneous cell populations.
Cell cytoplasm contains a complex mixture of chemicals and enzymes that are important for biochemical pathways and essential for the life of a cell. Abnormalities in the cytoplasm of subpopulations of cells are often a hallmark of pathology. While measurement of cytoplasmic components is essential to study cell physiology, most analysis methods only provide information regarding the cell population as a whole, not for individual cells or subpopulations of cells.
For example, only the overall concentration, but not the distribution of ATP concentrations in a heterogeneous cell population can be assessed using classical ATP measurements. ATP levels in cells can be sensitively measured using commercially available chemiluminescence assays. However, such analysis will only provide information of the cell population as a whole. In other words, a drop of ten percent (10%) in overall ATP concentration could be interpreted as ninety percent (90%) ATP remaining in all cells, or alternatively, that ten percent (10%) of the cells have no ATP at all. The consequences of a subpopulation of cells that is unable to maintain ATP seem obvious, but the determination of such a subpopulation is more difficult. Similar examples hold for many cytoplasmic components.
Single-cell analysis is thus an important tool in biology, as it defines properties of individual cells in heterogeneous cell populations. Current techniques, including flow cytometry, laser-scanning cytometry, and automated microscopy, are mainly used to define cell surface markers. Few probes are available that penetrate the plasma membrane and allow measurement of cytoplasmic components in single cells. Further, most cell analysis methods based on single-cell lysis are impractical to measure a significant number of cells in a reasonable amount of time to establish statistically relevant distributions in a cell population. The measurement of enzyme activity in single cells adds another complexity, as it requires monitoring of substrate use over time and under well-defined conditions.
In view of the foregoing, what are needed are apparatus and methods to analyze the stochastic distribution of cytoplasmic components in a heterogeneous cell population. Beneficially, such apparatus and methods would promote low cost, easy fabrication, high throughput, rapid measurement, and accurate correlation of cytoplasmic components with cell surface markers in a complex cell population. Such apparatus and methods are disclosed and claimed herein.